Objective: Prostate cancer is one of the most lethal forms of human cancer and paclitaxel is currently part of standard treatment for it. Combination therapy can enhance the anti-cancer activity of chemotherapy agents. Trifolium pratense L. has been suggested for cancer treatment in traditional medicine. In this study the effect of T. pratense hydroalcoholic extract on two prostate cell lines and normal fibroblast cells was investigated.

Materials and Methods: In this in vitro study, cells were treated with T. pratense extract and viability was evaluated using trypan blue staining, MTT assay, and lactate dehydrogenase activity measurement. Apoptosis and autophagy cell death were detected and quantified by fluorescent dyes staining. Nitric oxide production was measured using Griess reaction. Changes in expression level of some apoptotic and autophagic- related genes were investigated using Real-time PCR. The combination effect of extract and paclitaxel was evaluated by calculating the combination index and dose reduction index values.

Results: After treatment with T. pratense extract, the cell viability was significantly reduced in a time- and dose-dependent manner (p<0.05). The effect of extract on normal cells was significantly smaller than cancer cells (p>0.05). Apoptosis and autophagy cell death were significantly increased (p<0.05). Also, T. pratense extract significantly decreased NO production (p<0.05) by cancer cells. Combination of paclitaxel and T. pratense extract had a synergistic cytotoxic effect.

Conclusions: T. pratense showed an anti-cancer property via induction of apoptosis and autophagy cell death in prostate cancer cell lines.